Furan is an important industrial compound that is also present in the environment. It is both hepatoxic and carcinogenic in mice and rats. Based on these results and the potential for human exposure, furan has been listed as a possible human carcinogen. The mechanism of tumor induction by furan is unknown. The available experimental evidence suggests that both nongenotoxic and genotoxic mechanisms contribute to the overall carcinogenic outcome. It is clear that the toxicity and carcinogenicity is initiated by cytochrome P450 catalyzed oxidation of furan to an a,(3-unsaturated dialdehyde, c/s-2-butene-1,4-dial. However, it is possible that further metabolism of c/s-2-butene-1,4-dial generates other reactive metabolites that contribute to the toxic effects of furan. Our long-range goal is to determine the mechanism of furan-induced carcino- genesis in rodents so that we can develop effective tools to determine if humans exposed to furan are sus- ceptible to its toxicological properties. The objective of this application is to determine if furan is a genotoxic carcinogen in vivo and to better define the metabolic processes that lead to its activation or detoxification. The central hypothesis is that both cytotoxic and genotoxic pathways contribute to the overall carcinogenic effects of furan with the involvement of multiple metabolic pathways. We formulated this hypothesis on the basis of strong preliminary data that suggest that the oxidation of furan to c/s-2-butene-1,4-dial initiates the process. While this reactive metabolite can alkylate both DNA and protein in vitro, it is also likely to be further metabolized to other toxic compounds in vivo. We plan to test our hypothesis by pursuing the following specific aims: 1) Determine the mutagenic activity of furan in Big Blue rodents;2) Characterize the in vivo metabolic pathways of furan;3) Compare the pathways of furan metabolism in human and rodent hepatocytes and determine the enzymes involved in each pathway. Collectively, these specific aims will provide important data for testing mechanisms of furan toxicity and tumorigenesis. With this information, we can develop appropriate model systems to assess th6 role of genotoxicity and cytotoxicity in the carcino- genic properties of furan. In addition, these studies will aid in the development of appropriate biomarkers (urinary metabolites, protein and/or DNA adducts) that can be used to determine if these toxicologically important reactions are occurring in humans exposed to furan. This is a revised application from a well established investigator that seeks to further determine the mechanism of furan toxicity and mutagenicity. The strengths of this proposal are the strong background of the applicant in organic metabolite elucidation and DNA damage mechanisms, the clear and logical approach laid out in the specific aims and the responsiveness to the previous critiques. The weaknesses of this application are minimal. Some concerns are the potential for a null result in specific aims 1 and, particularly in aim 3 which may negate the use of a rodent model for determination of furan toxicity and mutagenicity in humans.